The two thiols of myosin, which when cross-linked by a variety of cross-linking agents trap MgADP at the active site, will be tagged with radioactive labels and the peptides containing these thiols isolated and identified by standard chromatographic and electrophoretic techniques. If appropriate, the peptides will be sequenced and compared with known sequences of peptides which contain the two activity-critical thiols of myosin. A new ATP photoaffinity label, 4-azido,2-nitroanilinoethyl tri-phosphate, will be trapped by cross-linking two thiols of myosin and subsequently covalently incorporated into myosin's active site by photolysis. The tryptic peptides containing the (32P)-labeled ATP analog will be isolated by gel chromatography and electrophoresis and compared to previous tryptic peptides isolated by others after labeling with a ATP analog containing a photolabile group attached to the ribose ring. The subunit location of the nucleotide binding will be determined by gel electrophoresis.